L-aminodicarboxylic acid esters

ABSTRACT

Sweeteners of the formula: ##STR1## and food-acceptable salts thereof, where the substituents are disclosed herein.

This invention is a continuation-in-part of copending U.S. applicationSer. No. 723,603 filed on Apr. 15, 1985, now abandoned.

This invention relates to a novel group of compounds and moreparticularly to a novel group of compounds particularly well suited assweeteners in edible foodstuff.

Sweetness is one of the primary taste cravings of both animals andhumans. Thus, the utilization of sweetening agents in foods in order tosatisfy this secondary desire is well established.

Naturally occuring carbohydrate sweeteners such as sucrose, are stillthe most widely used sweetening agents. While thse naturally occurringcarbohydrates, i.e., sugars, generally fulfill the requirements of sweettaste, the abundant usage thereof does not occur without deleteriousconsequence, e.g., high caloric intake and nutritional imbalance. Infact, oftentimes the level of these sweeteners required in foodstuffs isfar greater than the level of the sweetener that is desired foreconomic, dietetic or other functional consideration.

In an attempt to eliminate the disadvantages concomitant withcarbohydrate sweeteners, considerable research and expense have beendevoted to the production of artificial sweeteners, such as for example,saccharin, cyclamate, dihydrochalcone, aspatame, etc. While some ofthese artificial sweeteners satisfy the requirements of sweet tastewithout caloric input, and have met with considerable commercialsuccess, they are not, however, without their own inherentdisadvantages. For example, many of these artificial sweeteners have thedisadvantages of high cost, as well as delay in the perception of thesweet taste, persistent lingering of the sweet taste, and veryobjectionable bitter, metallic aftertaste when used in food products.

Since it is believed that many disadvantages of artificial sweeteners,particularly aftertaste, is a function of the concentration of thesweetener, it has been previously suggested that these effects could bereduced or eliminated by combining artificial sweeteners such assaccharin, with other ingredients such as aspartame or natural sugars,such as sorbitol, dextrose, maltose, etc. These combined products,however, have not been entirely satisfactory either. Some U.S. Patentswhich disclose sweetener mixtures include for example, U.S. Pat. No.4,228,198; U.S. Pat. No. 4,158,068; U.S. Pat. No. 4,154,862; and U.S.Pat. No. 3,717,477.

Accordingly, much work has continued in an attempt to develop andidentify compounds that have a sweet taste and which will satisfy theneed for better lower calorie sweeteners, and so research continues forsweeteners that have intense sweetness, that is, deliver a sweet tasteat low use levels and which will also produce enough sweetness at higherlevels to act as sole sweetener for most sweetener applications.Furthermore, the sweeteners sought must have good temporal and sensoryqualities. Sweeteners with good temporal qualities produce atime-intensity sweetness response similar to carbohydrate sweetenerswithout lingering. Sweeteners with good sensory qualities lackundesirable off tastes and aftertaste. Furthermore, these compounds mustbe economical and safe to use.

In U.S. Pat. No. 3,798,204, L-aspartyl-O-t-butyl-L-serine methyl esterand L-aspartyl-O-t-amyl-L-serine methyl ester are described as sweetcompounds having significant sweetness.

In U.S. Pat. No. 4,448,716 metal complex salts of dipeptide sweetenersare disclosed. In the background of this patent a generic formula isdescribed as an attempt to represent dipeptide sweeteners disclosed infive prior patents: U.S. Pat. No. 3,475,403; U.S. Pat. No. 3,492,131;Republic of South Africa Pat. No. 695,083 published July 10, 1969;Republic of South Africa Pat. No. 695,910 published Aug. 14, 1969; andGerman Pat. No. 2,054,554. The general formula attempting to representthese patents is as follows: ##STR2##

wherein R represents the lower alkyls, lower alkylaryls and cycloalkyls,n stands for integers 0 through 5, R₁ represents (a) phenyl group, (b)lower alkyls, (c) cycloalkyls, (d) R₂.

Where R₂ is hydroxy, lower alkoxy, lower alkyl, halogen, (e) (S(O)_(m)(lower alkyl) where m is 0, 1 or 2 and provided n is 1 or 2, (f) R₃.

Where R₃ represents hydroxy or alkoxy and (g) single or doubleunsaturated cycloalkyls with up to eight carbons. These compounds alsoare not entirely satisfactory in producing a high quality sweetness orin producing a sweet response at lower levels of sweetener.

Dipeptides of aspartyl-cysteine and aspartyl-methionine methyl estersare disclosed by Brussel, Peer and Van der Heijden in Chemical Sensesand Flavour, 4, 141-152 (1979) and in Z. Lebensm. Untersuch-Forsch.,159, 337-343 (1975). The authors disclose the following dipeptides:

α-L-Asp-L-Cys(Me)-OMe

α-L-Asp-L-Cys(Et)-OMe

α-L-Asp-L-Cys(Pr)-OMe

α-L-Asp-L-Cys(i-Pr)-OMe

α-L-Asp-L-Cys(t-But)-OMe

α-L-Asp-L-Met-OMe

In U.S. Pat. No. 4,399,163 to Brennan et al., sweeteners having thefollowing formulas are disclosed: ##STR3## and physiologicallyacceptable cationic and acid addition salts thereof wherein

R^(a) is CH₂ OH or CH₂ OCH₃ ;

R is a branched member selected from the group consisting of fenchyl,diisopropylcarbinyl, d-methyl-t-butylcarbinyl, d-ethyl-t-butyl-carbinyl,2-methylthio-2,4-dimethylpentan-3-yl, di-t-butyl-carbinyl, ##STR4##

In a related patent, U.S. Pat. No. 4,411,925, Brennan, et al. disclosecompounds of the above general formula with R being defined hereinabove,except R^(a) is defined as methyl, ethyl, n-propyl or isopropyl.

U.S. Pat. No. 4,375,430 to Sklavounos discloses dipeptide sweetenerswhich are aromatic sulfonic acid salts of L-aspartyl-D-alaninamides orL-aspartyl-D-serinamides.

European Patent Application No. 95772 to Tsau describe aspartyldipeptide sweeteners of the formula: ##STR5## wherein R' is alkyl of 1to 6 carbons, and R₂ is phenyl, phenylalkylenyl, or cyclohexylalkenyl,wherein the alkenyl group has 1 to 5 carbons. Closely related is U.S.Pat. No. 4,439,460 to Tsau, et al. which describes dipeptide sweetenersof the formula: ##STR6## wherein n is an integer from 0 to 5, and R₁ isan alkyl, alkylaryl or alicyclic radical. Similar such compounds aredescribed in many related patents, the major difference being thedefinition of R₂.

In U.S. Pat. No. 3,978,034 to Sheehan, et al., R₂ is defined ascycloalkenyl or phenyl. U.S. Pat. No. 3,695,898 to Hill defines R₂ as amono- or a di-unsaturated alicyclic radical. Haas, et al. in U.S. Pat.No. 4,029,701 define R₂ as phenyl, lower alkyl or substituted orunsubstituted cycloalkyl, cycloalkenyl or cycloalkadienyl, or S(O)_(m)lower alkyl provided that n is 1 or 2 and m is 0 or 2. Closely relatedare U.S. Pat. Nos. 4,448,716, 4,153,737, 4,031,258, 3,962,468,3,714,139, 3,642,491, and 3,795,746.

U.S. Pat. No. 3,803,223 to Mazur, et al. describe dipeptide sweetenersand anti-inflammatory agents having the formula: ##STR7## wherein R ishydrogen or a methyl radical and R' is a radical selected from the groupconsisting of alkyl, or ##STR8## wherein Alk is a lower alkyleneradical, X is hydrogen or hydroxyl, and Y is a radical selected from thegroup consisting of cyclohexyl, naphthyl, furyl, pyridyl, indolyl,phenyl and phenoxy.

Goldkamp, et al. in U.S. Pat. No. 4,011,260 describe sweeteners of theformula: ##STR9## wherein R is hydrogen or a lower alkyl radical, Alk isa lower alkylene radical and R' is a carbocyclic radical. Closelyrelated is U.S. Pat. No. 3,442,431.

U.S. Pat. No. 4,423,029 to Rizzi describes sweeteners of the formula:##STR10## Wherein R is C₄ -C₉ straight, branched or cyclic alkyl, andwherein carbons a, b and c have the (S) configuration.

European Patent Application No. 48,051 describes dipeptide sweeteners ofthe formula: ##STR11## wherein M represents hydrogen, ammonium, alkalior alkaline earth,

R represents ##STR12## R₁ represents methyl, ethyl, propyl, R₂represents --OH, or OCH₃,

* signifies an L-optical configuration for this atom.

German Patent Application No. 7259426 disclosesL-aspartyl-3-fenchylalanine methyl ester as a sweetening agent.

U.S. Pat. No. 3,971,822 to Chibata, et al., disclose sweeteners havingthe formula ##STR13## wherein R' is hydrogen or hydroxy, R₂ is alkyl ofone to five carbon atoms, alkenyl of two to three carbon atoms,cycloalkyl of three to five carbon atoms or methyl cycloalkyl of four tosix carbon atoms and Y is alkylene of one to four carbon atoms.

U.S. Pat. No. 3,907,366 to Fujino, et al. disclosesL-aspartyl-aminomalonic acid alkyl fenchyl diester and its'physiologically acceptable salts as useful sweeteners. U.S. Pat. No.3,959,245 disclose the 2-methyl cyclohexyl analog of the abovementionedpatent.

U.S. Pat. No. 3,920,626 discloses N-α L-aspartyl derivatives of loweralkyl esters of O-lower-alkanoyl-L-serine, β-alanine, γ-aminobutyricacid and D-β-aminobutyric acid as sweeteners.

Miyoshi, et al. in Bulletin of Chemical Society of Japan, 51, p.1433-1440 (1978) disclose compounds of the following formula assweeteners: ##STR14## wherein R' is H, CH₃, CO₂ CH₃, or benzyl and R₂ islower alkyl or unsubstituted or substituted cycloalkyl.

European Patent Application No. 128,654 describes gem-diaminoalkanesweeteners of the formula: ##STR15## wherein m is 0 or 1, R is loweralkyl (substituted or unsubstituted), R' is H or lower alkyl, and R" isa branched alkyl, alkylcycloalkyl, cycloalkyl, polycycloalkyl, phenyl,or alkyl-substituted phenyl, and physiologically acceptable saltsthereof.

U.S. Pat. No. 3,801,563 to Nakajima, et al. disclose sweeteners of theformula: ##STR16## wherein R' is a branched or cyclic alkyl group of 3to 8 carbon atoms, R₂ is a lower alkyl group of 1 to 2 carbon atoms andn is a integer of 0 or 1.

European Patent Application No. 34,876 describes amides ofL-aspartyl-D-amino acid dipeptides of the formula: ##STR17## whereinR^(a) is methyl, ethyl, n-propyl or isopropyl, and R is a branchedaliphatic, alicyclic or heterocyclic member which is branched at thealpha carbon atom and also branched again at one or both of the betacarbon atoms. These compounds are indicated to be of significantsweetness.

In the Journal of Medicinal Chemistry, 1984, Vol. 27, No. 12, pp.1663-8, are described various sweetener dipeptide esters, includingL-aspartyl-α-aminocycloalkane methyl esters.

The various dipeptide esters of the prior art have been characterized aslacking significant stability at low pH values and/or thermal stability.These characteristics have limited the scope of use of these sweetenersin food products which are of low pH values or are prepared or served atelevated temperatures.

Accordingly, it is desired to find compounds that provide qualitysweetness when added to foodstuffs or pharmaceuticals at low levels andthus eliminate or greatly diminish the aforesaid disadvantagesassociated with prior art sweeteners.

SUMMARY OF THE INVENTION

The present new compounds are esters of certain α-aminodicarboxylicacids and α-aminoesters which are low calorie sweeteners that possess ahigh order of sweetness with pleasing taste and higher stability at acidpH and elevated temperatures compared to known dipeptide sweeteners.

This invention provides new sweetening compounds represented by theformula: ##STR18## and food-acceptable salts thereof, wherein A ishydrogen, alkyl containing 1-3 carbon atoms, hydroxylalkyl containing1-3 carbon atoms or alkoxymethyl wherein the alkoxy contains 1-3 carbonatoms;

A' is hydrogen or alkyl containing 1-3 carbon atoms; alternatively;

A and A' taken together with the carbon atom to which they are attachedform cycloalkyl containing 3-4 carbon atoms;

Y is --(CHR₂)_(n) --R₁ or --CHR₃ R₄ ;

R₁ is an alkyl-substituted cycloalkyl, cycloalkenyl bicycloalkyl orbicycloalkenyl wherein at least one alkyl is in the β-position of thecycloalkyl, cycloalkenyl, bicycloalkyl or bicycloalkenyl ring,containing up to 7 ring carbon atoms and a total of 12 carbon atoms;

R₂ is H or alkyl containing 1-4 carbon atoms;

R₃ and R₄ are each cycloalkyl containing 3-4 ring carbon atoms;

n=0 or 1, and

m=0 or 1,

with the proviso that when the double asterisked carbon is an asymmetricor chiral center, the configuration around said carbon is in the D form.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

In accordance with the present invention, the preferred compounds arethose in which R₁ is an alkyl-substituted cycloalkyl or bicycloalkylcontaining 5-7 ring carbon atoms and up to a total of 10 carbon atoms.Especially preferred are cycloalkyl substituted with at least one methylgroup on the β and/or β' carbon atoms of the cycloalkyl ring.Particularly preferred cycloalkyls include cyclopropyl, cyclopentyl, andcyclohexyl and the preferred bicycloalkyl is fenchyl.

Also preferred are those compounds in which n=0. In those compounds inwhich n=1, R₁ is preferably a cyclopropyl group and R₂ is preferablytertiary butyl, isopropyl or cyclopropyl.

The groups representative of Y in the present new compounds include suchgroups as alkyl-substituted cycloalkyls, e.g., 1,2-dimethylcyclohexyl,1,2-dimethylcyclopentyl, 1,2-dimethylcycloheptyl,2,3-dimethylcyclopentyl, 2,3-dimethylcyclohexyl,2,3-dimethylcycloheptyl, 2,4-dimethylcyclopentyl,2,4-dimethylcyclohexyl, 2,4-dimethylcycloheptyl,2,5-dimethylcyclopentyl, 2,5-dimethylcyclohexyl,2,5-dimethylcycloheptyl, 2,6-dimethylcyclohexyl,2,6-dimethylcycloheptyl, 2,7-dimethylcycloheptyl,3,5-dimethylcyclopentyl, 4,5-dimethylcyclopentyl,4,5-dimethylcycloheptyl, 3,6-dimethylcyclohexyl,3,7-dimethylcycloheptyl, 4,6-dimethylcyclohexyl,4,7-dimethylcycloheptyl, 5,6-dimethylcyclohexyl, 5,6-dimethylcyclohexyl,5,7-dimethylcycloheptyl, 6,7-dimethylcycloheptyl,2,2-dimethylcyclopentyl, 2,2-dimethylcyclohexyl,2,2-dimethylcycloheptyl, 2,2,3-trimethylcyclopentyl,2,2,3-trimethylcyclohexyl, 2,2,3-trimethylcycloheptyl,2,2,4-trimethylcyclopentyl, 2,2,4-trimethylcyclohexyl,2,2,4-trimethylcycloheptyl, 2,2,5-trimethylcyclopentyl,2,2,5-trimethylcyclohexyl, 2,2,5-trimethylcycloheptyl,2,3,3-trimethylcyclopentyl, 2,3,3-trimethylcyclohexyl,2,3,3-trimethylcycloheptyl, 2,4,4-trimethylcyclopentyl,2,4,4-trimethylcyclohexyl, 2,4,4-trimethylcycloheptyl,1,2,3-trimethylcyclopentyl, 1,2,3-trimethylcyclohexyl,1,2,3-trimethylcycloheptyl, 1,2,4-trimethylcyclopentyl,1,2,4-trimethylcyclohexyl, 1,2,4-trimethylcycloheptyl,1,2,5-trimethylcyclopentyl, 1,2,5-trimethylcyclohexyl,1,2,5-trimethylcycloheptyl, 1,2,6-trimethylcyclohexyl,1,2,6-trimethylcycloheptyl, 1,2,7-trimethylcycloheptyl,2,3,4-trimethylcyclopentyl, 2,3,4-trimethylcyclohexyl,2,3,4-trimethylcycloheptyl, 2,3,5-trimethylcyclopentyl,2,3,5-trimethylcyclohexyl, 2,3,5-trimethylcycloheptyl,2,3,6-trimethylcyclohexyl, 2,3,6-trimethylcycloheptyl,2,3,7-trimethylcycloheptyl, 2,2,5,5-tetramethylcyclopentyl,2,2,5,5-tetramethylcyclohexyl, 2,2,5,5-tetramethylcycloheptyl,2,2,6,6-tetramethylcyclohexyl, 2,2,6,6-tetramethylcycloheptyl,2,2,7,7-tetramethylcycloheptyl, 2,2,4,4-tetramethylcyclopentyl,2,2,4,4-tetramethylcyclohexyl, 2,2,4,4-tetramethylcycloheptyl,2,2,3,3-tetramethylcyclopentyl, 2,2,3,3-tetramethylcyclohexyl,2,2,3,3-tetramethylcycloheptyl, 1,2,3,4-tetramethylcyclopentyl,1,2,3,4-tetramethylcyclohexyl, 1,2,3,4-tetramethylcycloheptyl,1,2,3,5-tetramethylcyclopentyl, 1,2,3,5-tetramethylcyclohexyl,1,2,3,5-tetramethylcycloheptyl, 1,2,3,6-tetramethylcyclohexyl,1,2,3,6-tetramethylcycloheptyl, 2,3,4,5-tetramethylcyclopentyl,2,3,4,5-tetramethylcyclohexyl, 2,3,4,5-tetramethylcycloheptyl,2,3,4,6-tetramethylcycloheptyl, 2,3,4,6-tetramethylcyclohexyl,2,3,4,7-tetramethylcycloheptyl, 2,2,3,4-tetramethylcyclopentyl,2,2,3,4-tetramethylcyclohexyl, 2,2,3,4-tetramethylcycloheptyl,2,2,3,5-tetramethylcyclopentyl, 2,2,3,5-tetramethylcyclohexyl,2,2,3,5-tetramethylcycloheptyl, 2,2,3,6-tetramethylcyclohexyl,2,2,3,6-tetramethylcycloheptyl, 2,2,3,7-tetramethylcycloheptyl,2,3,3,4-tetramethylcyclohexyl, 2,3,3,4-tetramethylcyclopentyl,2,3,3,4-tetramethylcycloheptyl, 2,3,3,5-tetramethylcyclopentyl,2,2,3,5-tetramethylcyclohexyl, 2,3,3,5-tetramethylcycloheptyl,2,3,3,6-tetramethylcyclohexyl, 2,3,3,6-tetramethylcycloheptyl,2,3,3,7-tetramethylcycloheptyl, 2,2,3,4-tetramethylcyclopentyl,2,2,3,4-tetramethylcyclohexyl, 2,3,3,4-tetramethylcycloheptyl,2,2,3,5-tetramethylcyclopentyl, 2,2,3,5-tetramethylcyclohexyl,2,2,3,6-tetramethylcyclohexyl, 2,2,3,6-tetramethylcycloheptyl,2,2,3,7-tetramethylcycloheptyl, 2,2,4,5-tetramethylcyclopentyl,2,2,4,5-tetramethylcyclohexyl, 2,2,4,5-tetramethylcycloheptyl,2,2,4,6-tetramethylcyclohexyl, 2,2,4,6-tetramethylcycloheptyl,2,2,4,7-tetramethylcycloheptyl, dicyclopropylmethyl,t-butylcyclopropylmethyl, dicyclobutylmethyl, t-butylcyclobutylmethyl,etc.; -alkyl-substituted cycloalkenes, e.g., 2-methyl-3-cyclohexenyl,2-methyl-3-cyclopentenyl, 2-methyl-3-cycloheptenyl,2-methyl-4-cycloheptenyl, 5-methyl-3-cyclopentenyl,2-methyl-2-cyclopentenyl, 2-methyl-2-cyclohexenyl,2-methyl-2-cycloheptenyl, 2-methyl-2-cyclopentenyl,6-methyl-2-cyclohexenyl, 7-methyl-2-cycloheptenyl,2,3-dimethyl-2-cyclopentenyl, 2,3-dimethyl-2-cyclohexenyl,2,4-dimethyl-2-cyclopentenyl, 2,4-dimethyl-2-cyclohexenyl,2,5-dimethyl-2-cyclohexenyl, 2,5-dimethyl-2-cycloheptenyl,2,6-dimethyl-2-cyclohexenyl, 2,6-dimethyl-3-cyclohexenyl,2,5-dimethyl-3-cyclohexenyl, 2,5-dimethyl-2-cyclopentenyl,2,4-dimethyl-3-cyclopentenyl, 2,4-dimethyl-3-cyclohexenyl,4,5-dimethylcyclo-3-pentenyl, 5,5-dimethyl-3-cyclopentenyl,6,6-dimethyl-3-cyclohexenyl, 1,2-dimethyl-3-cyclopentenyl,1,2-dimethyl-3-cyclohexenyl, 1,5-dimethyl-3-cyclopentenyl,2,2,6-trimethyl-3-cyclohexenyl, 2,2,5-trimethyl-3-cyclohexenyl,2,5,5-trimethyl-3-cyclohexenyl, 2,7,7-trimethyl-3-cycloheptenyl,2,7,7-trimethyl-4-cycloheptenyl, 2,2,7-trimethyl-3-cycloheptenyl,2,2,7-trimethyl-4-cycloheptenyl, 2,3,6-trimethyl-3-cyclohexenyl,2,3,7-trimethyl-3-cycloheptenyl, 2,3,5-trimethyl-3-cyclopentenyl,2,2,6,6 -tetramethyl-3-cyclohexenyl,2,2,5,5-tetramethyl-3-cyclopentenyl,2,2,7,7-tetramethyl-3-cycloheptenyl,2,3,5,5-tetramethyl-3-cyclopentenyl, 2,3,6,6-tetramethyl-3-cyclohexenyl,2,3,7,7-tetramethyl-3-cycloheptenyl,2,3,6,6-tetramethyl-3-cycloheptenyl, 2,3,5,5-tetramethyl-3-cyclohexenyl,2,3,4,5-tetramethyl-3-cyclopentenyl, 2,3,4,5-tetramethyl-3-cyclohexenyl,etc.; bicyclic compounds, such as norbornyl, norcaranyl, norpinanyl,bicyclo [2.2.2] octyl, etc.; alkyl substituted bicyclic compounds, e.g.,6,6-dimethyl-bicyclo [3.1.1] heptyl, 6,7,7-trimethylnorbornyl (bornyl orcamphanyl), pinanyl, thujanyl, caranyl, fenchyl, 2-norbornylmethyl,etc.; unsubstituted and alkyl-substituted bicycloalkenes such asnorbornenyl, norpinenyl, norcarenyl, 2-(4-norbornenyl)methyl, pinenyl,carenyl, fenchenyl, etc.; and tricyclo compounds such as adamantyl andalkyl-substituted adamantyl, etc.

The preferred R₁ is cycloalkyl or bicycloalkyl or alkyl-substitutedcycloalkyl or bicycloalkyl, especially where the alkyl group is in the βor β' positions. Further, preference exists for compounds in which R₁ isa cycloalkyl with two, three or four alkyl groups in the β, β' positionssuch as β, β, β', β'-tetraalkyl-substituted cyclopentyl, cyclobutyl,cyclohexyl, and cycloheptyl, as well as β, β, β'-trialkyl substitutedcyclobutyl, cyclopropyl, cyclohexyl, cyclopentyl, and cycloheptyl, andfenchyl. Also preferred are β-alkylcycloalkyls in which the alkyl groupis isopropyl or tertiary butyl.

These novel compounds are effective sweetness agents when used alone orin combination with other sweeteners in an ingesta, e.g., foodstuffs orpharmaceuticals. For example, other natural and/or artificial sweetenerswhich may be used with the novel compounds of the present inventioninclude sucrose, fructose, corn syrup solids, dextrose, xylitol,sorbitol, mannitol, acetosulfam, thaumatin, invert sugar, saccharin,thiophene saccharin, meta-aminobenzoic acid, metahydroxybenzoic acid,cyclamate, chlorosucrose, dihydrochalcone, hydrogenated glucose syrups,aspartame (L-aspartyl-L-phenylalanine methyl ester) and otherdipeptides, glycyrrhizin and stevioside and the like. These sweetenerswhen employed with the sweetness agents of the present invention, it isbelieved, could produce synergistic sweetness responses.

Furthermore, when the sweetness agents of the present invention areadded to ingesta, the sweetness agents may be added alone or withnontoxic carriers such as the abovementioned sweeteners or other foodingredients such as acidulants, natural and artificial gums, bulkingagents such as polycarbohydrates, dextrins, and other food approvedcarbohydrates and derivatives. Typical foodstuffs, and pharmaceuticalpreparations, in which the sweetness agents of the present invention maybe used are, for example, beverages including soft drinks, carbonatedbeverages, ready to mix beverages and the like, infused foods (e.g.vegetables or fruits), sauces, condiments, salad dressings, juices,syrups, desserts, including puddings, gelatin and frozen desserts, likeice creams, sherbets, icings and flavored frozen desserts on sticks,confections, chewing gum, cereals, baked goods, intermediate moisturefoods (e.g. dog food), toothpaste, mouthwash and the like.

In order to achieve the effects of the present invention, the compoundsdescribed herein are generally added to the food product at a levelwhich is effective to perceive sweetness in the food stuff and suitablyis in an amount in the range of from about 0.0005 to 2% by weight basedon the consumed product. Greater amounts are operable but not practical.Preferred amounts are in the range of from about 0.001 to about 1% ofthe foodstuff. Generally, the sweetening effect provided by the presentcompounds are experienced over a wide pH range, e.g. 2 to 10 preferably3 to 7 and in buffered and unbuffered formulations.

It is desired that when the sweetness agents of this invention areemployed alone or in combination with another sweetner, the sweetener orcombination of sweeteners provide a sucrose equivalent in the range offrom about 2 weight percent to about 40 weight percent and morepreferably from about 3 weight percent to about 15 weight percent in thefoodstuff or pharmaceutical.

A taste procedure for determination of sweetness merely involves thedetermination of sucrose equivalency. Sucrose equivalence for sweetenersare readily determined. The amount of a sweetener that is equivalent toa given weight percent sucrose can be determined by having a panel oftasters taste solutions of a sweetener at known concentrations and matchits sweetness to standard solutions of sucrose.

In order to prepare compounds of the present invention, several reactionschemes may be employed. In one reaction scheme, compounds of generalformula II (protected α-aminodicarboxylic acid) and III (amino-estercompound) are condensed to form compounds of general formula IV.Subsequent removal of protecting groups B and Z from compounds ofgeneral formula IV give the desired compounds of general formula I:##STR19## In these, group Z is an amino protecting group, B is acarboxyl protecting group, and A, A', Y, and n have the same meaning aspreviously described. A variety of protecting groups known in the artmay be employed. Examples of many of these possible groups may be foundin "Protective Groups in Organic Synthesis" by T. W. Green, John Wileyand Sons, 1981. Among the preferred groups that may be employed arebenzyloxycarbonyl for A and benzyl for B.

Coupling of compounds with general formula III to compounds havinggeneral formula IV employs established techniques in peptide chemistry.One such technique uses dicyclohexylcarbodiimide (DCC) as the couplingagent. The DCC method may be employed with or without additives such as4-dimethylaminopyridine or copper (II). The DCC coupling reactiongenerally proceeds at room temperature, however, it may be carried outfrom about -20° to 50° C. in a variety of solvents inert to thereactants. Thus suitable solvents include, but are not limited to,N,N-dimethyl-formamide, methylene chloride, toluene and the like.Preferably the reaction is carried out under an inert atmosphere such asargon or nitrogen. Coupling usually is complete within 2 hours but maytake as long as 24 hours depending on reactants.

Various other methods can be employed to prepare the desired compounds.The following illustrates such methods using aspartic acid as the aminodicarboxylic acid.

For example, U.S. Pat. Nos. 3,786,039; 3,833,553; 3,879,372 and3,933,781 disclose the reaction of N-protected aspartic anhydrides withamino acids and amino acid derivatives to yield the desired products.These N-protected aspartic anhydrides can be reacted with compounds offormula III by methods disclosed in the above patents. As described inU.S. Pat. No. 3,786,039 compounds of formula III can be reacted directlyin inert organic solvents with L-aspartic anhydride having its aminogroup protected by a formyl, carbobenzloxy, or p-methoxycarbobenzloxygroup which is subsequently removed after coupling to give compounds ofgeneral formula I. The N-acyl-L-aspartic anhydrides are prepared byreacting the corresponding acids with acetic anhydride in amounts of1.0-1.2 moles per mole of the N-acyl-L-aspartic acid at 0° to 60° C. inan inert solvent. The N-acyl-L-aspartic anhydrides are reacted withpreferably 1 to 2 moles of compounds of formula III in an organicsolvent capable of dissolving both and inert to the same. Suitablesolvents are, but not limited to, ethyl acetate, methyl propionate,tetrahydrofuran, dioxane, ethyl ether, N,N-dimethylformamide andbenzene. The reaction proceeds smoothly at 0° to 30° C. The N-acyl groupis removed after coupling by catalytic hydrogenation with palladium oncarbon or with HBr or HCl in a conventional manner. U.S. Pat. No.3,879,372 discloses that this coupling method can also be performed inan aqueous solvent at a temperature of -10° to 50° C. and at a pH of4-12.

Another method for the synthesis of the desired compounds is thereaction of compounds of formula III with suitable aspartic acidderivatives in which protecting groups have been attached to the aminoand beta-carboxy groups and the alpha carboxy group has been convertedto a reactive ester function. As disclosed in U.S. Pat. No. 3,475,403these coupled products may be deprotected as described to yield thedesired compounds of formula I.

An alternative scheme to the desired coupled compounds involves reactionof compounds of formula III with L-aspartic acid N-thiocarboxyanhydrideby the method of Vinick and Jung, Tet. Lett., 23, 1315-18 (1982). Anadditional coupling method is described by T. Miyazawa, Tet. Lett., 25,771 (1984).

Compounds of general formula III are synthesized using art recognizedtechniques. For example, compounds of formula III can be synthesized bystandard esterification methods known in the art by reacting the freeacid or acid functional equivalents, such as esters or anhydrides, withthe corresponding alcohols under ester-forming conditions, as forexample in the presence of mineral acids, such as hydrochloric orsulfuric acids or organic acids, such as p-toluene-sulfonic acids.Reaction temperatures are in the range of -78° to reflux. The reactionis carried out in a solvent that will dissolve both reactants and isinert to both as well. Solvents include, but are not limited tomethylene chloride, diethyl ether, tetrahydrofuran, dimethylsulfoxide,N,N-dimethylformamide, and the like.

With regard to the removal of protecting groups from compounds offormula IV and N-protected precursors of formula III, a number ofdeprotecting techniques are known in the art and can be utilized toadvantage depending on the nature of the protecting groups. Among suchtechniques is catalytic hydrogenation utilizing palladium on carbon ortransfer hydrogenation with 1,4-cyclohexadiene. Generally the reactionis carried at room temperature but may be conducted from 5° to 65° C.Usually the reaction is carried out in the presence of a suitablesolvent which may include, but are not limited to water, methanol,ethanol, dioxane, tetrahydrofuran, acetic acid, t-butyl alcohol,isopropanol or mixtures thereof. The reaction is usually run at apositive hydrogen pressure of 50 psi but can be conducted over the rangeof 20 to 250 psi. Reactions are generally quantitative taking 1 to 24hours for completion.

In any of the previous synthetic methods the desired products arepreferably recovered from reaction mixtures by crystallization.Alternatively, normal or reverse-phase chromatography may be utilized aswell as liquid/liquid extraction or other means.

The desired compounds of formula I are usually obtained in the free acidform; they may also be recovered as their physiologically acceptablesalts, i.e., the corresponding amino salts such as hydrochloride,sulfate, hydrosulfate, nitrate, hydrobromide, hydroiodide, phosphate orhydrophosphate; or the alkali metal salts such as the sodium, potassium,lithium, or the alkaline earth metal salts such as calcium or magnesium,as well as aluminum, zinc and like salts.

Conversion of the free peptide derivatives of formula I into theirphysiologically acceptable salts is carried out by conventional means,as for example, bringing the compounds of formula I into contact with amineral acid, an alkali metal hydroxide, an alkali metal oxide orcarbonate or an alkaline earth metal hydroxide, oxide, carbonate orother complexed form.

These physiologically acceptable salts can also be utilized as sweetnessagents usually having increased solubility and stability over their freeforms.

It is known to those skilled in the art that the compounds of thepresent invention having asymmetric carbon atoms may exist in racemic oroptically active forms. All of these forms are contemplated within thescope of the invention.

The compounds of the present invention have one asymmetric site, whichis designated by an asterisk (*) in the formula below, and onepseudoasymmetric site which is designated by a double asterisk (**).##STR20## Furthermore, depending upon the substituent, Y may alsocontain chiral centers. All of the stereochemical configurations areencompassed within the above formula. However, the present invention isdirected to compounds of the formula: ##STR21## Although both D and Lforms are possible, the present invention is directed to those compoundsin which the dicarboxylic acid group is in the L-configuration asdepicted in Formula I.

Whenever A is identical to A', or A and A' together form anunsubstituted cyclopropyl or cyclobutyl group, the compounds of thepresent invention have at least one asymmetric site, designated by theasterisk in the dicarboxylic acid moiety.

Whenever the group A and A' are different, the carbon atom designated bythe double asterisk become as asymmetric center and a chiral center andthe compounds of Formula I will contain at least two asymmetric centers.Furthermore, when A and A' taken together form a cyclopropyl orcyclobutyl ring having substituents, said carbon atom designated by thedouble asterisk may have an asymmetric center. In those cases whereinthe carbon atom designated by the double asterisk is a chiral center,Formula I encompasses compounds of Formula II having the L, Lconfiguration and Formula III having the L, D configuration: ##STR22##In the instance wherein the carbon designated by the double asterisk isa chiral center, the preferred compounds are those in which theconfiguration around the double asterisked carbon is in the Dconfiguration. In the production of compounds of Formula I, the L, Ldiastereomer though not sweet itself, may be admixed with the L, Dstereoisomers. The admixture of the L, L and L, D stereoisomers exhibitsweetness, but said mixture is not as sweet as the compound of FormulaIII (i.e., the L, D stereoisomer) in its pure form.

The following examples further illustrate the invention. In thefollowing examples, the sensory evaluation were obtained by a panel ofexperts using known weight percent aqueous solutions of the exemplifiedcompounds and were matched to sucrose standard solutions.

EXAMPLE 1 L-Aspartyl-D-alanine(2,2,5,5-tetramethyl-1-cyclopentyl)ester

To a magnetically stirred solution of 10 g (0.071 mol)2,2,5,5-tetramethylcyclopentanone in 75 ml of dry tetrahydrofuran at 0°C. under argon is added 2.69 g (0.071 mol) of lithium aluminum hydride.When the reduction was complete, ethyl acetate was introduced dropwiseto destroy unreacted lithium aluminum hydride. 25 mls of water is thenadded, followed by 300 mls of diethyl ether. The organic phase is washedwith 100 mls of water, and dried over MgSO₄. Filtration followed byevaporation afforded 8.23 g of 2,2,5,5-tetramethylcyclopentanol.

To a stirred solution of 9.59 g (0.043 mol) N-Cbz-alanine in 50 ml dryCH₂ Cl₂ containing 8.96 g (1 eq.) dicyclohexylcarbodiimide and 0.4 gdimethylaminopyridine (DMAP), all at 0° C., is added, via an additionfunnel, 6.0 g (0.043 mol) of 2,2,5,5-tetramethylcyclopentanol dissolvedin 50 ml CH₂ Cl₂. After stirring for 48 hours, the mixture is filtered,and the filtrate is washed with 5% HCl (1×50 ml), saturated NaHCO₃ (1×50ml), and water (1×50 ml). The organic layer is separated, dried overMgSO₄ and evaporated to yield 5.68 g of crude material, which aftersilica gel chromatography yielded 5.68 g of2,2,5,5-tetramethylcyclopentyl N-Cbz-D-alanine ester.

5.68 g of 2,2,5,5-Tetramethylcyclopentyl N-Cbz-D-alanine ester isdissolved in 100 ml CH₃ OH and hydrogenated over 5% Pd/C in a Parrhydrogenation apparatus. When the reaction is complete the mixture isfiltered through Celite and concentrated to yield 3.75 g of2,2,5,5-tetramethylcyclopentyl alanine ester.

To a magnetically stirred solution of 3.75 g (0.017 mol)2,2,5,5-tetramethylcyclopentyl alanine ester in 170 ml of drydimethylformamide at 0° C. under atgon atmosphere is added 6.07 g (0.017mol) N-Cbz-L-aspartic acid beta-benzyl ester followed by 2.28 g copper(II) chloride, and 3.54 g dicyclohexylcarbodiimide. This is stirred for18 hours, after which the reaction mixture is poured into 200 ml 0.1NHCl and extracted with 300 ml ethyl acetate. The organic phase is washedwith saturated NaHCO₃ and then water, and dried over MgSO₄. Evaporationof the solvent followed by silica gel chromatography yielded 5.14 gN-(N'-Cbz-L-aspartyl beta-benzyl ester)-D-alanine2,2,5,5-tetramethyl-1-cyclopentyl ester.

2.0 g N-(N'-Cbz-L-aspartyl beta-benzyl ester)-D-alanine2,2,5,5-tetramethyl-1-cyclopentyl ester is dissolved in 50 ml CH₃ OH andhydrogenated over 5% Pd/C in a Paar apparatus. Upon completion of thereaction th mixture is filtered and concentrated to yield 2.59 gL-aspartyl-D-alanine 2,2,5,5-tetramethyl-1-lcyclopentyl ester. [α]_(D)²⁵ (pure)=+21.9°.

NMR (DMSO): δ0.8 (s, 6H), 1.00 (s, 6H), 1.3 (d, 3H), 1.45 (s, 4H),2.2-2.4 (m, 2H), 4.35 (s, 1H), 4.75 (bris).

FAB-MS (m/z): 329 (M-H, 22%), 205 (20%), 90 (17%), 69 (100%).

Sweetness determination with this compound gave the following results:

    ______________________________________                                                         SUCROSE                                                      CONCENTRATION    EQUIVALENCE                                                  ______________________________________                                        0.005            2.5                                                          0.010            3.7                                                          0.025            7.7                                                          0.05             8.0                                                          ______________________________________                                    

Similarly, by utilizing the appropriate alcohol, the followingadditional compounds are prepared:

N-L-Aspartyl-D-alanine(2,2,5-trimethylcyclopentyl)ester.

N-L-Aspartyl-D-alanine(2,5-dimethylcyclopentyl)ester.

N-L-Aspartyl-D-alanine(dicyclopropylmethyl)ester.

N-L-Aspartyl-D-alanine(fenchyl)ester.

N-L-Aspartyl-D-alanine(2-t-butylcyclopentyl)ester.

N-L-Aspartyl-D-alanine(1-t-butyl-1-cyclopropylmethyl)ester.

N-L-Aspartyl-D-alanine(1-isopropyl-1-cyclopropylmethyl)ester.

EXAMPLE 2N-(L-Aspartyl)-2-methylalanine(2,2,5,5-tetramethyl-1-cyclopentyl)ester

To a stirred solution of 2 g (0.008 mol) of N-Cbz-2-aminoisobutyric acidin dry Cl(CH₂)₂ Cl containing 1.9 g dicyclohexylcarbodiimide and 0.1 gdimethylaminopyridine (DMAP), all at 0° C., is added, via an additionfunnel, 1.3 g of 2,2,5,5-tetramethylcyclopentanol dissolved in CH₂ Cl₂.After stirring for 4 days, the mixture is filtered, and the filtrate iswashed with 5% HCl (1×50 ml), saturated NaHCO₃ (1×50 ml), and water(1×50 ml). The organic layer is separated, dried over MgSO₄ andevaporated to yield N-Cbz-2-aminoisobutyric acid2,2,5,5-tetramethylcyclopentyl ester.

N-Cbz-2-aminoisobutyric acid 2,2,5,5-tetramethylcyclopentyl ester isdissolved in CH₃ OH and hydrogenated over 10% Pd/C in a Paarhydrogenation apparatus. When the reaction is complete the mixture isfiltered through Celite and concentrated to yield 2-aminoisobutyric acid2,2,5,5-tetramethylcyclopentyl ester (600 mg).

To a magnetically stirred solution of 0.6 g 2-aminoisobutyric acid2,2,5,5-tetramethylcyclopentyl ester in 20 ml of dry dimethylformamideat 0° C. under argon atomosphere was added 1.02 g N-Cbz-L-aspartic acidbeta-benzyl ester followed by 0.38 g of copper (II) chloride and 0.58 gdicyclohexylcarbodiimide. This is stirred for 18 hours, after which thereaction mixture is poured into 0.1N HCl and extracted with ethylacetate. The organic phase is washed with saturated NaHCO₃ and thenwater, and dried over MgSO₄. Evaporation of the solvent yieldsN-(N'-Cbz-L-Aspartyl beta-benzylester)-2-methylalanine(2,2,5,5-tetramethyl-1-cyclopentyl)ester.

N-(N'-Cbz-L-Aspartyl beta-benzylester)-2-methylalanine(2,2,5,5-tetra-methyl-1-cyclopentyl)ester isdissolved in CH₃ OH and hydrogenated over 5% Pd/C in a Parr apparatus.Upon completion of the reaction the mixture is filtered and concentratedto yield crudeN-(L-Aspartyl)-2-methylalanine(2,2,5,5-tetramethyl-1-cyclopentyl)ester.Purification of the final product was done by reverse-phasechromatography on a Whatman Magnum 20 ODS-3 C₁₈ column; solvent system:75% methanol in H₂ O.

NMR (CDCl₃ /DMSO): δ0.90 (s, 6H), 1.00 (s, 6H), 1.25-1.5 (m, 4H), 1.50(s, 6H), 2.50-2.70 (q. of d., 2H), 3.50, 3.85 (br.s, 1H), 4.85 (br.s).

Sweetness determination with this compound gave the following results:

    ______________________________________                                                         SUCROSE                                                      CONCENTRATION    EQUIVALENCE                                                  ______________________________________                                        0.005            2.00                                                         0.010            3.25                                                         0.025            5.75                                                         ______________________________________                                    

Similarly, by utilizing the appropriate alcohol, the followingadditional compounds are prepared:

N-L-Aspartyl 2-methylalanine(2,2,5,-trimethylcyclopentyl)ester.

N-L-Aspartyl 2-methylalanine(2,5-dimethylcyclopentyl)ester.

N-L-Aspartyl 2-methylalanine(dicyclopropylmethyl)ester.

N-L-Aspartyl 2-methylalanine(fenchyl)ester.

N-L-Aspartyl 2-methylalanine(2-t-butylcyclopentyl)ester.

N-L-Aspartyl 2-methylalanine(1-t-butyl-1-cyclopropylmethyl)ester.

N-L-Aspartyl 2-methylalanine(1-isopropyl-1-cyclopropylmethyl)ester.

EXAMPLE 3 N-(L-Aspartyl)-1-amino-1-cyclopropanecarboxylic acid(2,2,5,5-tetramethyl-1-cyclopentyl)ester

To a stirred solution of N-Cbz-1-aminocyclopropane carboxylic acid indry (CH₂)₂ Cl₂ containing dicyclohexylcarbodiimide anddimethylaminopyridine (DMAP), all at 0° C., is added, via an additionfunnel, 2,2,5,5-tetramethylcyclopentanol dissolved in CH₂ Cl₂. Afterstirring for 4 days, the mixture is filtered, and the filtrate is washedwith 5% HCl (1×50 ml), saturated NaHCO₃ (1×50 ml), and water (1×50 ml).The organic layer is separated, dried over MgSO₄ and evaporated to yieldN-Cbz-1-aminocyclopropanecarboxylic acid 2,2,5,5-tetramethylcyclopentylester.

N-Cbz-1-aminocyclopropanecarboxylic acid 2,2,5,5-tetramethylcyclopentylester is dissolved in absolute alcohol at 0° C. in an ultrasound bath.Palladium on carbon (10%) is added. The hydrogen source,1,4-cyclohexadiene, is added, and ultrasound is commenced for eightminutes. The slurry is then filtered through a bed of Celite with ethylalcohol. The solvent is removed by rotary evaporation to yield1-aminocyclopropylcarboxylic acid 2,2,5,5-tetramethylcyclopentyl ester.

To a magnetically stirred solution of 1-aminocyclopropane carboxylicacid 2,2,5,5-tetramethylcyclopentyl ester is dry dimethylformamide at 0°C. under argon atmosphere is added N-Cbz-L-aspartic acid beta-benzylester followed by copper (II) chloride and dicyclohexylcarbodiimide.This is stirred for 18 hours, after which the reaction mixture is pouredinto 0.1N HCl and extracted with ethyl acetate. The organic phase iswashed with saturated NaHCO₃ and then water, and dried over MgSO₄.Evaporation of the solvent yields N-(N'-Cbz-L-Aspartyl beta-benzylester)-1-amino-1-cyclopropanecarboxylic acid2,2,5,5-tetramethyl-1-cyclopentyl ester.

The N-(N'-Cbz-L-Aspartyl-beta-benzylester)-1-amino-1-cyclopropanecarboxylic acid2,2,5,5-tetramethyl-1-cyclopentyl ester is dissolved in absolute alcoholat 0° C. in an ultrasound bath. Palladium on carbon (10%) is added. Thehydrogen source, 1,4-cyclohexadiene, is added, and ultrasound iscommenced for eight minutes. The slurry is then filtered through a bedof Celite with ethyl alcohol. The solvent is removed by rotaryevaporation to afford the final product.

Similarly, by utilizing the appropriate starting materials the followingadditional compounds are prepared:

N-L-aspartyl 1-aminocyclopropane-1-carboxylicacid(2,2,5,-trimethylcyclopentyl)ester.

N-L-aspartyl 1-aminocyclopropane-1-carboxylicacid(2,5-dimethylcyclopentyl)ester.

N-L-aspartyl 1-aminocyclopropane-1-carboxylic acid(dicyclopropylmethyl)ester.

N-L-aspartyl 1-aminocyclopropane-1-carboxylic acid (fenchyl)ester.

N-L-aspartyl 1-aminocyclopropane-1-carboxylic acid(2-t-butylcyclopentyl)ester.

N-L-aspartyl 1-aminocyclopropane-1-carboxylic acid(1-t-butyl-1-cyclopropylmethyl)ester.

N-L-aspartyl 1-aminocyclopropane-1-carboxylic acid(1-isopropyl-1-cyclopropylmethyl)ester.

The sweetness determination withL-Aspartyl-1-aminocyclopropyl-1-carboxylic acid,2,5-dimethyl-1-cyclopentyl ester gave the following results:

    ______________________________________                                                                  Sweetness                                                                     relative to                                                                   Sucrose                                             % Compond   Sucrose Equivalents                                                                         (× Sucrose)                                   ______________________________________                                        0.005       1.0           200                                                 0.010       2.2           217                                                 0.025       3.3           133                                                 ______________________________________                                    

EXAMPLE 4N-L-Aspartyl-O-methyl-D-serine(2,2,5,-trimethylcyclopentyl)ester

To a solution of 5 g N-Cbz-D-serine 2,2,5-trimethylcyclopentyl ester in50 ml dry CH₂ Cl₂ is added 2 equivalents of Ag₂ O and 2 equivalents ofmethyl iodide. After stirring for 2 hours, the mixture is filtered andconcentrated to yield the methyl ether of N-Cbz-D-serine2,2,5-trimethylcyclopentyl ester. 3 g of N-Cbz-D-serine methyl ether2,2,5-trimethylcyclopentyl ester is hydrogenated over 0.5 g 10% Pd/C in100 ml CH₃ OH. Upon completion, the mixture is filtered and concentratedto yield 3-methoxy-D-alanine 2,2,5-trimethylcyclopentyl ester. To amagnetically stirred solution of 2 g of 3-methoxy-D-alanine2,2,5-trimethylcyclopentyl ester in 100 ml DMF at 0° C. is added 1equivalent of N-Cbz-L-aspartic acid-β-benzyl ester followed by additionof 1 equivalent each of Cu(II) chloride and dicyclohexylcarbodiimide.After 18 hours the mixture is poured into 200 ml 0.1N HCl and extractedwith 300 ml ethyl acetate. The organic phase is washed with saturatedNaHCO₃, and H₂ O, dried over MgSO₄, filtered and concentrated to an oilthat is reconstituted in 50 ml CH₃ OH and hydrogenated over 0.5 g 5%Pd/C. Filtration followed by concentration yields L-aspartyl-D-serine2,2,5-trimethylcyclopentyl ester methyl ether.

Using the appropriate starting materials, the following dipeptides areadditionally prepared:

N-L-Aspartyl-O-methyl-D-serine(2,5-dimethylcyclopentyl)ester.

N-L-Aspartyl-O-methyl-D-serine(dicyclopropylmethyl)ester.

N-L-Aspartyl-O-methyl-D-serine(fenchyl)ester.

N-L-Aspartyl-O-methyl-D-serine(2-t-butylcyclopentyl)ester.

N-L-Aspartyl-O-methyl-D-serine(1-t-butyl-1-cyclopropylmethyl)ester.

N-L-Aspartyl-O-methyl-D-serine(1-isopropyl-1-cyclopropylmethyl)ester.

N-L-Aspartyl-O-methyl-D-serine(2,2,5,5-tetramethylcyclopentyl)ester.

EXAMPLE 5 L-Aspartyl-D-serine-(2,2,5-trimethylcyclopentyl)ester

Into a suspension of N-Cbz-D-serine (5 g) in 50 ml of dry THF containing1 equivalent of 2,2,5-trimethylcyclopentanol is bubbled dry hydrogenchloride gas at room temperature. Upon complete solution of the mixture,the reaction is refluxed for 5 hours, then concentrated. Ethyl acetateis added, and this is washed with saturated sodium bicarbonate, water,and dried over MgSO₄. Filtration followed by concentration yieldsN-Cbz-D-serine-2,2,5-trimethylcyclopentyl ester. 5 g of this product isdissolved in 10 ml methanol and hydrogenated in a Paar apparatus over 1g of 5% Pd/C to yield 2,2,5-trimethylcyclopentyl-D-serinate.

To a magnetically stirred solution of 0.1 mole2,2,5-trimethylcyclopentyl-D-serinate in 100 ml dry DMF at 0° C. underan argon atmosphere is added 1 equivalent of N-Cbz-L-aspartic acidβ-benzyl ester followed by addition of 1 equivalent each of Cu(II)chloride and dicyclohexylcarbodiimide. After 18 hours the mixture ispoured into 200 ml 0.1N HCl and extracted with 300 ml ethyl acetatewhich is successively washed with saturated NaHCO₃, H₂ O, and dried overMgSO₄. Filtration and evaporation yieldsN-Cbz-β-benzyl-L-aspartyl-D-serine 2,2,5-trimethylcyclopentyl ester. 2 gN-Cbz-β-benzyl-L-aspartyl-D-serine 2,2,5-trimethylcyclopentyl ester in50 ml dry CH₃ OH is hydrogenated in a Paar apparatus over 5% Pd/C. Uponcompletion of the reaction, the mixture is filtered through Celite andconcentrated to dryness to yield the final product.

Similarly, utilizing the appropriate starting materials the followingadditional compounds are prepared:

N-L-Aspartyl-D-serine(2,2,5-trimethylcyclopentyl)ester.

N-L-Aspartyl-D-serine(2,5-dimethylcyclopentyl)ester.

N-L-Aspartyl-D-serine(dicyclopropylmethyl)ester.

N-L-Aspartyl-D-serine(fenchyl)ester.

N-L-Aspartyl-D-serine(2-t-butylcyclopentyl)ester.

N-L-Aspartyl-D-serine(1-t-butyl-1-cyclopropylmethyl)ester.

N-L-Aspartyl-D-serine(1-isopropyl-1-cyclopropylmethyl)ester.

N-L-Aspartyl-D-serine(2,2,5,5-tetramethylcyclopentyl)ester.

EXAMPLE 6 N-L-Aspartyl-D-alanine(1-methyl-1-cyclopentyl)ester A.N-carbobenzoxy-D-alanine(1-methyl-1-cyclopentyl)ester

To a magnetically stirred solution of 22.3 g (0.1 mol) N-Cbz-D-alaninein 50 mls of dry dichloromethane containing 0.5 mls of concentratedsulfuric acid at 0° C., was added dropwise a 10 g (0.1 mol) sample of1-methylcyclopentene in 50 mls of dichloromethane. After 5 days ofstirring at room temperature, the mixture was heated to reflux for 4hours, after which the reaction was cooled to room temperature, washedwith 100 mls of saturated NaHCO₃, 100 mls of water and dried over MgSO₄.Filtration followed by evaporation of the solvent yielded 1.81 g of theproduct. NMR(CDCl₃): δ1.3-1.4(d, 3H), 1.5(s, 3H), 1.5-1.7 (m, 8H), 4.2(m, 1H), 5.05 (s, 2H), 5.25 (m, 1H), 7.3 (s, 5H).

B. D-Alanine(1-methylcyclopentyl)ester

1.8 g of the compound of part A was hydrogenated in 50 mls of methanolcontaining 0.5 g of 5% Pd/C catalyst in a Paar apparatus. The catalystwas filtered off, the solvent was removed by evaporation and 0.54 g of1-methylcyclopentyl D-alanine ester was obtained.

C.Beta-benzyl-N-carbobenzoxy-L-aspartyl-D-alanine-(1-methyl-1-cyclopentyl)ester

To 0.54 g (0.0031 mol) of the product from B in 31 mls ofdimethylformamide at 0° C. under an argon atmosphere is added 1.11 g(0.0031 mol) of N-Cbz-L-aspartic acid, betabenzyl ester, followed by 417mg (1 equiv.) Cu (II) Cl₂ and 646 mg. (1 equiv.)dicyclohexylcarbodiimide. This is stirred for 16 hours, after which itis poured into 200 mls of 0.1N HCl and extracted with 3×100 ml of ethylacetate. The organic phase was washed with 100 ml of water and driedover MgSO₄. Filtration and evaporation of the solvent yielded 1.0 g ofbeta-benzyl-N-carbobenzoxyL-aspartyl-D-alanine-(1-methyl-1-cyclopentyl)ester.

NMR (CDCl₃): δ1.3-1.4(d, 3H), 1.5 (s, 3H), 1.5-1.7 (m, 8H), 2.7-3.0 (dof d, 2H), 4.35 (m, 2H), 5.1 (s, 2H), 5.8 (d, 1H), 6.9 (d, 1H), 7.3 (s,5H).

D. N-L-aspartyl-D-alanine(1-methyl-1-cyclopentyl)ester

2.3 g of the product from C was hydrogenated over 0.5 g of Pd/C (5%) inmethanol to yield 280 mg.L-aspartyl-D-alanine(1-methyl-1-cyclopentyl)ester.

NMR (D₂ O): δ1.3-1.4(d, 3H), 1.5 (s, 3H), 1.5-1.7 (m, 8H), 2.3 (m, 2H),4.2 (m, 2H).

Sweetness determination with this compound gave the following results:

    ______________________________________                                                                 Sweetness Value                                                    Sucrose    Relative to                                          Percent of Compound                                                                         Equivalence                                                                              Sucrose (× Sucrose)                            ______________________________________                                        0.05          2.0        40                                                   ______________________________________                                    

EXAMPLE 7 N-L-Aspartyl-D-alanine(2,5-dimethylcyclopentyl)ester A.N-carbobenzoxy-D-alanine(2,5-dimethylcyclopentyl)ester

To a magnetically stirred solution of 19.63 g (0.008 mol)N-Cbz-D-alanine, 18.34 g (1 equiv.) of dicyclohexylcarbodiimide, and0.88 g of 4-(dimethylamino)pyridine in 150 mls of dichloromethane at 0°C., is added 10 g (0.088 mol) of 2,5-dimethylcyclopentanol. After 48hours, the reaction mixture is filtered to remove dicyclohexylurea andconcentrated to a pale yellow oil, which is redissolved in ethylacetate. This is successively washed with 100 mls of 5% HCl, 100 mls ofsaturated NaHCO₃, 100 mls of saturated NaCl and 100 mls of water, driedover MgSO₄ and filtered. Evaporation of the solvent afforded 23.7 ofN-Cbz-D-alanine, 2,5-dimethylcyclopentylester.

B. D-Alanine-(2,5-dimethylcyclopentyl)ester

5.08 g of the product from A was hydrogenated several times over 0.5 gof 5% Pd/C in 50 mls of CH₃ OH to yield 2.59 g ofD-alanine-(2,5-dimethylcyclopentyl)ester.

C. N-Cbz-beta-benzyl-L-aspartyl-D-alanine(2,5-dimethylcyclopentyl)ester

To a solution of 9.45 g (0.05 mol) of the product from B in 300 mls ofdry DMF at 0° C. is added 17.85 g (1 eq.) ofN-Cbz-beta-benzyl-L-aspartic acid, 6.72 g (1 eq.) copper (II) chlorideand 10.42 g (1 eq.) dicyclohexylcarbodiimide. This, at 0° C. under anargon atmosphere, is stirred for 18 hours. The mixture is filtered toremove the urea, and is poured onto 300 mls of 0.1N HCl. The bluesolution is extracted with 3×200 ml of diethyl ether and 3×200 ml ofethyl acetate. The combined organic phases were washed with 100 mls ofNaHCO₃ (saturated), 100 mls of saturated NaCl, and 100 mls of water, anddried over MgSO₄. Filtration and evaporation afforded 29.6 g of thecrude above-identified product, which was purified by flash silica gelchromatography.

D. N-L-Aspartyl-D-alanine(2,5-dimethylcyclopentyl)ester

The product from C was hydrogenated in the usual fashion in methanolover 5% Pd/C to yield the final product. [α]_(D) ²⁵ =2.7°.

Sweetness determination with this compound gave the following results:

    ______________________________________                                                                  Sweetness relative                                                            to Sucrose                                          Concentration                                                                            Sucrose Equivalence                                                                          (× Sucrose)                                   ______________________________________                                        0.005      1.16           233                                                 0.01       2.3            230                                                 0.025      4.82           193                                                 0.050      7.50           150                                                 ______________________________________                                    

EXAMPLE 8 N-L-Aspartyl-1-amiocyclopropane carboxylic acid(2,2,5,5-tetramethyl-1-cyclopentyl)ester A.N-t-butoxycarbonyl-1-aminocyclopropanecarboxylic acid (1)

To a solution of 1-aminocyclopropanecarboxylic acid (3.03 g) insaturated aqeuous sodium bicarbonate (150 ml) was added a solution ofdi-t-butyldicarbonate (9.82 g) in t-butanol (50 ml), and the resultingmixture was stirred overnight. Water was then added and the mixture waswashed with ethyl acetate. The aqueous phase was separated, made acid topH 1 with concentrated hydrochloric acid and extracted twice with ethylacetate. The combined extracts were washed with saturated sodiumchloride, dried over magnesium sulfate, and the solvent was evaporatedto yield a white solid. (4.77 g, 86%). NMR (CDCl₃): δ1.05-1.35 (m, 2H,cyclopropyl), 1.41 (s, 9H, t-butyl), 1.41-1.70 (m, 2H, cyclopropyl),5.20, (br. s, 1H, NH), 9.25 (br. s, 1H, CO₂ H).

B. 2,5-DimethylcyclopentylN-t-butoxycarbonyl-1-aminocyclopropanecarboxylate (2)

To a solution of 2,5-dimethylcyclopentanol (0.55 g.), compound 1 (0.97g), and 4-(dimethylamino)pyridine (0.06 g.) in methylene chloride (100ml) was added dicyclohexylacarbodiimide (1.09 g.), and the resultingmixture was stirred overnight. The precipitated dicyclohexylurea wasremoved by filtration, and the filtrate was evaporated. Ethyl acetatewas then added to the residue, and the mixture was filtered again. Thefiltrate was washed with 1M hydrochloric acid, saturated aqeuous sodiumbicarbonate, and water, dried over magnesium sulfate, and the solventwas evaporated to a colorless oil (1.18 g. 83%). The product waspurified by column chromotography on silica gel, 4:1 hexane:ethylacetate, eluent.

NMR (CDCl₃): δ0.92 (t, 6H, 2CH₃), 1.05-2.10 (m, 10H, cyclopentyl,cyclopropyl), 1.40 (s, 9H, t-Bu), 4.60 (dd., 1H, CO₂ CH), 5.05 (br. s,1H, NH).

C. β-Benzyl-N-benzyloxycarbonyl-L-aspartyl-1-aminocyclopropanecarboxylicacid, 2,5-dimethylcyclopentyl ester (3)

A mixture of compound 2 (0.57 g), 95% ethanol (11 ml), water (7.5 mls)and concentrated hydrochloric acid (4 ml) was heated to reflux for 2hours. The mixture was cooled, and 1 molar hydrochloric acid was added.The solution was washed with ethyl acetate. The separated aqueous phasewas made basic with 1M sodium hydroxide and extracted twice with ethylacetate. The combined extracts were washed with saturated sodiumchloride, dried over magnesium sulfate, and the solvent was evaporatedto yield 0.22 g of a colorless oil. Dicyclohexylcarbodiimide (0.25 g)was added to a solution of the above oil, followed byN-benzyloxycarbonyl-L-aspartic acid, beta-benzyl ester (0.40 g) andcopper (II) chloride (0.17 g) in dimethyl formamide (10 mls). Theresulting mixture was stirred overnight. The green mixture was thenfiltered to remove dicyclohexylurea, and 1M hydrochloric acid was addedto the filtrate. This was extracted twice with ethyl acetate, and thecombined extracts were washed with 1M hydrochloric acid, saturatedsodium bicarbonate and saturated sodium chloride. The solution was driedover magnesium sulfate, and the solvent was evaporated to give 0.53 g ofa yellow oil. This is purified by column chromotography on silica gel,4:1 hexane:ethyl acetate, eluent, to give the desired product as a whitesolid (0.33 g, 32%). NMR (CDCl₃): δ0.91 (t, 6H, 2CH₃), 0,80-2.25 (m,10H, cyclopropyl, cyclopentyl), 2.70 (dd., 1H, J₁ =7 Hz, J₂ =17 Hz,aspartyl CH₂), 3.00 (dd., 1H, J₁ =5 Hz, J₂ =17 Hz, aspartyl CH₂),4.30-4.70 (m, 2H, CO₂ CH), aspartyl CH), 5.10 (s, 4H, 2PhCH₂), 5.87 (br.d, 1H, NH), 6.95 (br. s, 1H, NH), 7.30 (s, 10H, 2Ph).

D. L-Aspartyl 1-aminocyclopropanecarboxylic acid,2,5-dimethylcyclopentyl ester

A mixture of compound 3 (0.31 g), 1,4-cyclohexadiene (0.46 g), 10%palladium-on-carbon (0.3 g) and 95% ethanol (10 ml) was placed in anultrasonic bath for 10 minutes. The mixture was then filtered throughCelite, and the solvent was evaporated to yield 130 mg of a colorlessoil which solidified upon standing. This was purified by HPLC using areverse phase C₁₈ column, 60% methanol in water as eluent to yield thedesired product as a white solid (72 mg, 40%). MP: 155.5°-157° C. FAB MS(m/z): 313 (M+H, 22%) 217 (87%), 102 (69%), 88 (100%).

EXAMPLE 9 α-L-Aspartyl-2-methylalanine [β(+)Fenchyl]ester

N-CBZ-protected amino isobutyric acid (Chemical Dynamics, Inc.) wasdissolved in 1,2-dichloroethane (50 mL) at 0° C. under argon. A solutionof N,N-dimethylaminopyridine (0.5 equiv.) and β(+) fenchyl alcohol (1equiv.) in 1,2-dichloroethane (10 mL) was added. Lastly,dicyclohexylcarbodiimide (1.1 equiv.) was added as a solid. After fivedays of stirring at room temperature the urea was removed by filtrationand the filtrate was diluted with petroleum ether (50 mL). The solutionwas clarified again by filtration and the filtrate was hi-vacuum rotaryevaporated to a paste. Column chromatography on silica gel with 15:1petroleum ether/ethyl acetate gave the pure product in 75-79% yield as awhite crystalline solid. NMR (CDCl₃): δ0.90 (s, 3H), 1.05 (s, 3H) 1.10(s, 3H), 1.20-1.80 (m, 7H), 1.60 (s, 6H), 4.20 (s, 1H), 5.10 (s, 2H),5.55 (s, 1H), 7.40 (s, 5H). [α]_(D) ²⁵ =-11.65° (MeOH) mp. 83°-85° C.

The ester from above was deprotected in the usual manner byhydrogenation with palladium on carbon (10%) in methanol to give aquantitative yield of the free-amino ester.

The amine was immediately dissolved in DMF and coupled to an asparticacid precursor by the Copper (II) chloride procedure to give a 90% yieldof N-CBZα-L-aspartic acidβ-benzylesterα2-methylalanine[β(+)Fenchyl]ester. NMR (CDCl₃): δ0.90 (s,3H), 1.05 (s, 3H), 1.10 (s, 3H), 1.20-1.80 (m, 7H), 1.6 (d, 6H)2.70-3.15 (m, 2H), 4.1-4.2 (m, 1H), 4.20 (s, 1H), 4.60 (s, 1H), 5.10 (s,4H), 5.60 (d, 1H), 5.90 (d, 1H), 5.90 (d, 1H), 7.40 (s, 10H). Theproduct was deprotected by hydrogenation and purified by Rp C₁₈ columnchromatography with 85:15 methanol:water eluant, [α]_(D) ²⁵ =-3.30°(MeOH) mp. 121°-3° C.

Sweetness determination with this compound gave the following results:

    ______________________________________                                                                  Sweetness relative to                               Concentration                                                                           Sucrose Equivalence                                                                           Sucrose (× Sucrose)                           ______________________________________                                        0.00750   8.5%            1133                                                0.00375   6.0%            1600                                                0.00185   5.7%            3100                                                0.00692   3.5%            3800                                                ______________________________________                                    

EXAMPLE 10 α-L-Aspartyl-D-alanine [β(+) Fenchyl]ester A.exo-β-(+)-Fenchol

To a refluxing suspension of 72.65 g aluminum isopropoxide in 300 ml offreshly distilled isopropyl alcohol, was added dropwise, 27.1 gR-(-)-fenchone in 50 ml isopropanol. The reaction was halted after sixdays when it was determined by gas chromatography (Carbowax 20M) thatmore than 50% of the ketone was reduced. It was also determined bycapillary chromatography (Supelcowax 10) that the exo/endo ratio for thefenchol was 3/1. Upon cooling, the mixture was filtered and washedthoroughly with dichloromethane. The precipitate was dissolved in 5% HCl(100 ml) and extracted with dichloromethane (50 ml). The combineddichloromethane solutions were washed with 5% HCl (50 ml), saturatedNaHCO₃ (50 ml) and water (50 ml) and dried over MgSO₄. Filtration andremoval of the solvent afforded 23.44 g of an oil that was 40% unreactedfenchone and 60% α and β fenchol isomers.

A mixture of 12 g (0.78 mol) β and α-fenchols, 11.9 ml (1.1 eq)triethylamine and 15.9 g p-nitrobenzol chloride (1.1 g) in 500 mls drydichlormethane was refluxed for 24 hours. The mixture ofβ/α esters wasseparated by silica gel flash chromatography using hexane:ethyl acetate(40:1). 6.0 g of the exo-fenchyl para-nitrobenzate was isolated,([α]_(D) ²⁵ =-17.1° (in benzene). 3 g of fenchol (9/1; β/α) was obtainedupon basic hydrolysis of the nitrobenzate ester (refluxing excess NaOHin methanol). β-(+)-fenchol; [α]_(D) ²⁵ =+23.4° (neat),

NMR: δ0.95-1.8 (16H, m, CH₂, CH₃); 3.0 ppm (1H, s, CH--O).

B. N-Cbz-D-alanine, β-(+)-fenchyl ester

To a stirred solution of 1.3 g β-(+)-fenchol in 20 ml drydichloromethane was added 1.9 g (0.0084 mol) N-Cbz D-alanine, and thesolution was cooled to 0° C. Then, 0.113 g p-dimethylaminopyridine and1.91 g dicyclohexylcarabodiimide were added. After 24 hours, thereaction was stopped and filtered. The solvent was evaporated and theoily residue was dissolved in diethyl ether, washed with 5% HCl (25 ml),saturated NaHCO₃ (25 ml), water (25 ml) and dried over MgSO₄. Afterfiltration and solvent evaporation, the product was purified by silicagel chromatography to yield 1.86 g N-Cbz-D-alanine, β-(+)-fenchyl ester;[α]_(D) ²⁵ =+3.86°.

NMR: δ0.8-1.8 ppm (19H, m, CH₂, CH₃); 4.2 ppm (1H, s, CH--O); ##STR23##5.1 ppm (2H, s, CH₂ --Ph) 5.4 ppm (1H, d, NH); 7.4 ppm (5H, s, Ph).

C. D-alanine, β(+)-fenchyl ester

The N-Cbz-D-alanine, β-(+)-fenchyl ester, (1.86 g) was dissolved in 50ml methanol and hydrogenated over 0.1 g 5% Pd/C in a Paar shaker. After2 hours the reaction was over; it was filtered through Celite, washedwith methanol, concentrated and the crystallized residue was dissolvedin dichloromethane.

D. N-Cbz-β-benzyl-L-aspartyl-D-alanine, β-(+)fenchyl ester

To the DMF solution containing the D-alanine ester (0.00355 mol) wasadded an equimolar amount of B-benzyl-N-Cbz-L-aspartic acid (1.27 g) and0.526 g Cu(II)Cl₂. Upon solution of the CuCl₂, DCC (0.81 g) was added.After 24 hours, the reaction was complete, the urea was filtered and thesolvent was evaporated. The yellow oil was dissolved in diethyl ether(25 ml) and washed with 5% HCl (25 ml), saturated NaHCO₃ (25 ml), and H₂O (25 ml). The ether layer was dried over MgSO₄ and evaporated to yield0.95 g of product. NMR: δ0.85-1.80 (19H, m, CH₂, CH₃), 4.2 ppm (1H, s,CH--O); ##STR24## 5.1 ppm (4H, s, OCH₂ --Ph); 5.95 ppm (1H, d, NH); 7.05ppm (1H, d, NH); 7.4 ppm (10H, s, Ph).

E. L-aspartyl-D-alanine, β-(+)-fenchyl ester

0.95 g protected dipeptide was dissolved in 50 ml methanol to which 0.1g 10% Pd/C was added. This was hydrogenated in Paar shaker for 2 hours.The solution was filtered and evaporated to dryness to yield 0.194 gsolid; [α]_(D) ²⁵ =-0.867°.

The product was purified on reverse phase HPLC (85% methanol/water) toyield 75 mg L-aspartyl-D-alanine, β-(+)Fenchyl ester. NMR: δ0.8-1.8(19H, m, CH₂, CH₃); ##STR25## 4.2 ppm (1H, s, OCH); 4.5 ppm (2H, m,N--CH); ##STR26##

Sweetness determination with this compound gave the following results:

    ______________________________________                                                                  Sweetness relative to                               Concentration                                                                           Sucrose Equivalence                                                                           Sucrose (× Sucrose)                           ______________________________________                                        0.00750   9.0%            1200                                                0.00375   8.6%            2300                                                0.00185   6.5%            3500                                                0.00092   4.7%            5100                                                ______________________________________                                    

The compounds of this invention possess greater sweetness and higherstability in comparison to corresponding esters of the prior art.

What is claimed is:
 1. N-L-Aspartyl-D-alanine [β(+)fenchyl]ester. 2.N-L-Aspartyl-2-methylalanine [β(+)Fenchyl]ester.